This application represents an interdisciplinary effort to critically evaluate and improve on current methodologies for the design and analysis of surface plasmon resonance (SPR) experiments as they are used to investigate protein:protein interactions. SPR offers a powerful technique to determine the kinetics of macromolecular interactions, the experimental set-up often implies that the measured data deviates from that predicted based on the underlying kinetic model. These deviations can lead to incorrect analysis of the experimental data. The deviations can arise from a multitude of sources, such as mass transport and rebinding effects. It is currently recommended to run kinetic experiments under conditions that result in a low signal to noise ratio which can lead to further difficulties in the data analysis. It is hypothesized that the application of techniques which have been developed in the signal processing literature can resolve some of the existing problems.
The aims of this application are to: 1) to investigate the effects of reference cell subtraction and baseline drift on the measured kinetic constants and to develop methods that overcome the potential problems; 2) to develop methods to improve on the currently available techniques to deal with mass transport, rebinding, low signal to noise ratios and fast off rates; 3) to develop further methods for the identification of the correction interaction model for a given protein:protein interaction; and 4) to develop a software environment in which these studies can be efficiently carried out.
